20.109(F11):Katie Vogel and Tonia Tsinman

From OpenWetWare

Our Idea
It's well known that most of the water on our planet is stored in the form of salt water found in oceans. Plants and animals, however, can only survive by using a fresh water source, which is a much smaller percentage of the total water supply. Hence, recent research has focused on increasing the tolerance of plant cells to salinity. A design of an effective technique to accomplish such a goal holds great promise, as this would mean that we could potentially re-engineer common crops such that they could be watered using salt water! Moreover, high soil salt concentration remains one of the primary reasons for why land cannot be used for agricultural purposes. Thus, genetically engineering plants to grow in places with high salt concentration holds great promise for increased productivity and efficiency of the agricultural industry.

Thus far, several approached to re-engineering plant cells have been taken. One path has been to screen plant germplasts in vitro for salinity tolerance and mate more tolerant plants together to produce offspring that are increasingly better at growing in water with increasing concentrations of salt.
This research proposal focuses on utilizing the second approach, namely the creation of mutant cell lines and the generation of plant cells from the desired mutants. Specifically, we want to use the approach we applied to the bacterial photography system optimization, and set up a mutant screen to look for decreased uptake of sodium ions in plant cells.

In order to accomplish this, the pathway of sodium uptake at the roots of cells is needed and is shown below:

Given the mechanisms of the pathway, two options could potentially limit Na+ input.
1. Inhibition of the HKT1 Na+ transporter. We would have to do more research on the structure of this transporter, but more research needs to be done to examine which residues are key to the transporter function. Using site directed mutagenesis, the transporter can be blocked and uptake of Na+ could be limited.
[1]

2. Altering the kinasing activity of Salt overly sensitive(SOS) transduction pathway. SOS3 activates SOS2, a kinase responsible for activating the SOS1 plasma membrane pump responsible for expelling Na+. Research could be done to increase the kinasing activity of SOS2 and examine if the plant cell could tolerate higher salinity with a more active Na+ pump. [2]

Predictions and Problems

If everything is completely successful, the inactivation of one of these should cause reduced uptake of Na+, making cells more saline tolerant.
However, altering HKT1 could lead to problems with the cell maintaining proper turgor pressure at all times, even when the cell is not in a state of saline stress. SOS2 regulates NHX1, which is responsible for vacuole sodium balance. Modifying SOS2 could possibly effect osmotic regulation of the vacuole. Also, unlike two component signaling systems, which are responsible for saline resistance in rice, modifying a signaling cascade could cause unknown adverse effects that we cannot be predicted due to the complexity of the signaling cascade.